PROJECT SUMMARY/ABSTRACT Regenerative capacity is widespread throughout almost all animal phyla, but the distributing pattern appears to be inexplicable. This diverse regenerative distribution raises questions of how animals evolve toward loss or gain of regenerative capacity and of what cellular and molecular mechanisms control regenerative ability. One feature of regeneration is that innervation is essential for peripheral tissue regeneration. Previous studies have shown that nerves are involved in multiple regeneration processes from early to late regenerative events and that distinct neuronal subtypes, such as cholinergic and sensory neurons, play different roles during tissue regeneration. However, whether neuronal excitation is required for tissue regeneration and which neuronal subtypes are associated with these processes remains poorly known. Obtaining a genetically amenable animal model will uniquely permit the identification of the essential neuronal subtypes and establishing their roles in tissue regeneration. Through forward genetic screening to discover novel regeneration-associated genes, we recently discovered a new zebrafish mutant exhibiting locomotion disorder and impaired fin regeneration in a temperature-dependent manner. Whole-exome sequencing and further fine genetic mapping analysis identified a missense mutation in the scn8a gene, which encodes the major neuronal voltage-gated sodium channel Nav1.6. We will develop a new paradigm for ion channel-regulated tissue regeneration. We will take advantage of the temperature sensitivity of scn8a mutation to define how scn8a mutation influences locomotion behavior and multiple regenerative processes. We will elucidate principles for neurons as essential drivers of tissue regeneration. We will address the challenge of whether neuronal Scn8a is required for locomotion and tissue regeneration and which Scn8a expressing neuronal subtypes are associated with fin regenerative processes. In addition to scn8a mutant, we will investigate the other two mutants, each of which exhibits either fin re-growth defects or impaired re-patterning, to uncover unidentified regeneration-associated genes and underlying mechanisms. The proposed study will construct genetic models for tissue regeneration, leading to the discovery of valuable genes regulating tissue regeneration and establishment of regenerative networks.